Nerve growth factor (NGF) is a polypeptide required for the survival and development of sympathetic, sensory, and certain central nervous system neurons. It binds to specific cell surface receptors on these neurons, intiates a chain of intracellular events, and, through these intracellular actions, controls the expression of specific genes. The molecular mechanisms by which the factor controls gene expression are not known, but, through the work in this and other laboratories, the cascade of intracellular events mediating the actions of the factor are becoming clear. The binding of nerve growth factor to its receptor on the NGF-responsive cell line, PC12, is followed rapidly by an activation of phosphoinositide metabolism and an activation protein kinase C. This activation appears due to a phosphorylation of the protein Kinase C. Kinase C and other kinases lead to changes in the phosphorylation of a number of key proteins in the cell with resultant changes in their activity. Among these are two proteins involved in protein synthesis, the S6 protein of the ribosomes and the elongation factor 2 (EF-2) in the cytoplasm; also altered is the phosphorylation of a nuclear protein (SMP), perhaps proximally involved in gene transcription. We have developed cell-free systems for the phosphorylation of these three proteins in order to dissect the biochemical mechanisms by which the phosphorylations are altered. A major effort now is to find that single reaction coupling the receptor to these kinase pathways. This will be aided by the finding, in this laboratory, of an inhibitor, K-252a, that is specific for the actions of nerve growth factor. It has been shown, for example, that the induction of certain oncogenes, notably c-fos, by nerve growth factor is prevented by K-252a, but induction of c-fos by other ligands is unaffected. Alterations in other gene products are also being explored in order to find out which gene products and in which order are required for the alterations in differentiation produced by nerve growth factor. We have found that nerve growth factor causes the appearance of a characteristic neuronal marker, binding sites for tetanus toxin, that must require the expression of certain glycoprotein genes. We have also found that nerve growth factor treatment causes a decrease in the number of receptors for epidermal growth factor, a mitogen for these cells. An understanding of the mechanism by which the differentiating agent causes a decrease in the receptors for the mitogen could provide an important insight into the overall control of differentiation and cell division.